Uso di tecnologie omiche per lo sviluppo di farmaci: studio su composti 3-cloropiperidinici

Tumor heterogeneity and complex resistance mechanisms undermine the therapeutic value of existing anticancer options, generating a pressing demand for alternative accessible and effective leads. In this sense, our group recently introduced 3-chloropiperidines (3-CePs) as a novel class of alkylating agents designed to improve the pharmacological profile of nitrogen mustard (NM) chemotherapeutics. Previous works described the improved alkylating properties of bifunctional 3-chloropiperidines (B-CePs) compared to the reference NM chlorambucil, exploring the effect of different linker structures on their reactivity with DNA and dissecting the mechanism of B-CePs alkylation at guanines. From these premises, this project further investigated the chemical space and antiproliferative value of these anticancer candidates. First, an explorative set of monofunctional analogues (M-CePs) was introduced to study the effect of a single reactive center on the activity of compounds. Notably, despite reduced DNA cleavage relative to the bifunctional compounds, these new agents showed nanomolar cytotoxicity indexes against test cancer cells, along with an unexpected tropism for a pancreatic adenocarcinoma cell line (BxPC-3), demonstrating to be valid alternatives for the future development of this class of compounds. Previous works showed that aromatic linkers decreased the alkylation potency compared to aliphatic analogues. Since classical nitrogen mustards with milder electrophilicity are known to be also less toxic in vivo, we further expanded our library of compounds with a new and enlarged set of aromatic B-CePs. Interestingly, aromatic agents exerted a remarkable activity against 2D and 3D cancer cell cultures as well as an exclusive tropism for BxPC-3 cells, as observed for M-CePs. As a general rule, slower reactivity and enhanced transported-mediated uptake profiles were demonstrated to boost cytotoxicity against the more complex model of three-dimensional spheroids. The tropism for pancreatic adenocarcinoma BxPC-3 cells was further observed for a third set of Lys-ester derivatives bearing appended instead of embedded aromatic linkers. In this other study, an extended set of derivatives bearing Lys-ester/amide linkers with various aromatic and aliphatic side chains was investigated in terms of cellular activity and consumption kinetics due to ring hydroxylation in aqueous solution, a competing reaction which abates the fraction of compound able to alkylate the biological target. Our results highlighted that the lower DNA reactivity of Lys-ester aromatic analogues was compensated by a longer stability to hydroxylation, allowing them to preserve valuable indexes of cytotoxicity. In the last work presented here, we investigated the molecular bases of the observed 3-CePs tropism for the pancreatic adenocarcinoma cell line. Representative mono- and bifunctional compounds were thoroughly analyzed by the use of a multi-omic approach to investigate the molecular determinants of cell susceptibility to our drug candidates. We analyzed transcriptional changes and chromatin status upon treatment in a high- (pancreatic adenocarcinoma BxPC-3) and low-sensitive (colorectal adenocarcinoma HCT-15) cancer cell lines, demonstrating that BxPC-3 cells were unable to control proteostasis and DNA damage response in stress conditions when exposed to our alkylating agents. Furthermore, we derived perturbation-informed signatures predicting compound sensitivity and identified potentially more susceptible target tumor types for the further development of clinical candidates. We believe that our results exemplify the potential of a multi-omic approach in offering a versatile framework to support drug discovery toward precision oncology.